The reduction of the m6A methyltransferase METTL3 in granulosa cells is related to the follicular cysts in pigs.

Follicular cysts are a common reproductive disorder in domestic animals that cause considerable economic losses to the farming industry. Effective prevention and treatment methods are lacking because neither the pathogenesis nor formation mechanisms of follicular cysts are well-understood. In this study, we first investigated the granulosa cells (GCs) of cystic follicles isolated from pigs. We observed a significant reduction in the expression of methyltransferase-like 3 (METTL3). Subsequent experiments revealed that METTL3 downregulation in GCs caused a decrease in m6A modification of pri-miR-21. This reduction further inhibited DGCR8 recognition and binding to pri-miR-21, dampening the synthesis of mature miR-21-5p. Additionally, the decrease in miR-21-5p promotes IL-1ß expression in GCs. Elevated IL-1ß activates the NFκB pathway, in turn upregulating apoptotic genes TNFa and BAX/BCL2. The subsequent apoptosis of GCs and inhibition of autophagy causes downregulation of CYP19A1 expression. These processes lower oestrogen secretion and contribute to follicular cyst formation. In conclusion, our findings provide a foundation for understanding and further exploring the mechanisms of follicular-cyst development in farm animals. This work has important implications for treating ovarian disorders in livestock and could potentially be extended to humans.

Abundance of Anti-Muellerian hormone in cat ovaries and correlation of its plasma concentration with animal age, weight and stage of the estrous cycle.

In female animals of different species, Anti-Müllerian hormone (AMH) is produced by follicular granulosa cells and has been associated with the ovarian follicle pool. Because concentration of AMH in plasma of ovary-intact female cats is apparently more variable than previously assumed, we have analysed AMH concentration in blood of cats (n = 93) presented for routine ovariectomy and assessed ovarian histology and AMH protein expression in the surgically removed ovaries. We hypothesised that AMH is synthesized only in preantral and small antral follicles and that plasma AMH concentration reflects the antral follicle count (AFC). Corpora lutea were detected in 35% of the female cats, whereas plasma progesterone concentration was ≥1 ng/mL in 57% of the cats. Follicular cysts were present in 15 cats (16%). Positive immunostaining for AMH protein was detected in close to all primordial and antral follicles, ovarian cysts, 70% of corpora lutea and 28% of atretic follicles. Concentration of AMH in plasma averaged 6.8 ± 0.5 ng/mL (range 1.3-21.7 ng/mL). The AFC increased with increasing AMH concentration with a moderate positive correlation between AFC and AMH (r = 0.286, p < 0.01). Plasma AMH concentration was not affected by season or cats' age, weight, stage of the estrous cycle and presence of follicular cysts. In conclusion, AMH protein is expressed in all endocrine structures of the cat ovary. While AMH is a marker for the presence of ovarian tissue, its usefulness to assess ovarian function in individual female cats is of limited value.

A review on inflammation and angiogenesis as key mechanisms involved in the pathogenesis of bovine cystic ovarian disease.

Cystic ovarian disease (COD) is an important cause of reproductive failure in dairy cattle. The main aim of this review is to discuss some aspects related to inflammation and angiogenesis that seem to be involved in the development of follicular cysts in domestic animals, with special emphasis on the bovine species, in an attempt to elucidate the relationship between these two processes in the early stages of persistence and in the development of bovine COD. We describe the changes in the expression of cytokines and angiogenic factors that seem to generate disturbances in the intraovarian component underlying the aberrant persistence of follicular cysts. Results show that pro-inflammatory and anti-inflammatory cytokines behave as regulators of angiogenesis through direct and indirect effects, like overexpression of pro-angiogenic factors, particularly in bovine ovarian cells from follicular cysts and persistent follicles. We conclude that, in dairy cattle, an imbalance in the expression of cytokines and pro-angiogenic growth factors related to ovulation and the processes associated with it would contribute to follicular persistence and to the recurrent appearance of COD.

Altered vitamin D metabolic system in follicular cysts of sows.

This study aimed to examine 25OHD3 concentration in the fluid of follicular and follicular lutein cysts of sows in comparison with preovulatory follicles as well as immunolocalize vitamin D metabolic enzymes (CYP27B1 and CYP24A1) and determine their protein abundances in the cyst wall. We have shown for the first time that 25OHD3 level in the fluid of both cyst types was significantly lower than in preovulatory follicles. Furthermore, we have demonstrated CYP27B1 and CYP24A1 protein immunolocalization and abundance in follicular and follicular lutein cysts. The abundance of protein for both metabolic enzymes was decreased in ovarian cysts when compared to preovulatory follicles. We propose that altered VD metabolism in ovarian cyst might associate with their formation in sows.

Contribution of the VEGF system to the follicular persistence associated with bovine cystic ovaries.

Cystic ovaries (CO) characterize a disorder frequently found in dairy cattle. However, despite the contributions by several researchers, the mechanism that leads to ovulatory failure has not yet been completely elucidated. Thus, the aim of this study was to examine the mRNA expression of bovine vascular endothelial growth factor (VEGFA)-164, VEGFA-164b and VEGF receptors (VEGFR1 and VEGFR2) by real-time PCR and protein expression by immunohistochemistry, immunofluorescence and Western blot in follicular fluid from dairy cows with spontaneous CO and in an experimental model of follicular persistence induced by prolonged treatment with progesterone. Results showed that both VEGFA isoforms and receptors were coexpressed in granulosa and theca interna cells and in follicular fluid of ovaries from all the groups evaluated. VEGFA-164, VEGFA-164b and VEGFR2 protein expression was higher in theca cells of persistent follicles from group P0 (expected time of ovulation) than in those from dominant follicles (as reference structure) from the control group (p < 0.05). Also, VEGFA-164 expression was higher in theca cells of cysts than in those of dominant follicles of the control group (p < 0.05). In follicular fluid, VEGFA-164 expression was higher in persistent follicles from group P5 (5 days of follicular persistence) than in the control, P0 and P15 groups, and higher in cysts than in dominant follicles from the control group (p < 0.05). This study provides evidence of an altered expression of VEGFA-164, VEGFA-164b and VEGFR2 during the formation of persistent follicles and cysts in cows. Together, these results evidence that early development of CO in cows is concurrent with an altered expression of these growth factors and that these alterations may contribute to the follicular persistence, angiogenic dysregulation and ovulatory failure found in cows with follicular cysts.

Alterations in key metabolic sensors involved in bovine cystic ovarian disease.

High-producing dairy cows frequently suffer metabolic alterations that cause different diseases, which could decrease the reproductive efficiency of the herd. Among these reproductive disorders, cystic ovarian disease (COD) has been related to alterations in metabolites and hormonal factors such as insulin, adiponectin and leptin. The aim of this study was to determine the protein expression of adiponectin and some of its downstream targets in ovarian follicles of control cows and cows with clinical diagnosis of COD. We also analyzed some key metabolic sensors in plasma and follicular fluid from both groups. In follicular cysts, we detected higher protein expression of adiponectin receptor 2 (AdipoR2), 5' adenosine monophosphate-activated protein kinase (AMPK), carnitine palmitoyl transferase 1 (CPT1) and acyl-coenzyme A oxidase 1 (ACOX1) relative to control antral follicles (p < 0.05). This was related to higher plasma adiponectin concentration in cows with COD than in control cows (p < 0.05). On the other hand, insulin concentrations showed an opposite pattern (p < 0.05). Furthermore, we found alterations in local and systemic concentrations of several metabolites. In this regard, in follicular fluid of cystic cows, the concentrations of non-esterified fatty acids and beta-hydroxybutyrate were higher (p < 0.05), whereas the concentrations of glucose and triacylglycerol were lower than in follicular fluid from control cows (p < 0.05). Besides, in both follicular fluid and plasma of cows with COD, the concentration of cholesterol was higher than in control animals (p < 0.05). These results evidence a local altered scenario of some metabolic sensors in cystic follicles, which could generate an adverse microenvironment for the resumption of ovarian activity, possibly causing the persistence of follicles and the recurrence of COD.

Follicular Cysts: A Single Sign and Different Diseases. A View from Comparative Medicine.

Ovarian cystic follicles are the sign of important causes of reproductive failure in numerous species. In this review, some morphological, endocrinological and clinical aspects of cystic follicles in women, cows, mares, sows and bitches are discussed. Follicular cysts are the consequence of the failure of a mature follicle to ovulate at the appointed time of ovulation in the estrous cycle. Although the etiology of follicular cysts remains unknown, this review examines the evidence about the role of endocrine signaling systems in the specific disease or syndrome in each of the species mentioned above. This review also describes, the changes in the pathways of endocrine mechanisms that would trigger disturbances in the intraovarian component underlying the aberrant persistence of follicular cysts. The knowledge of the morphological and endocrinological nature of cystic follicles in different species can provide relevant information to better understand specific diseases when it is integrally analyzed from the comparative medicine viewpoint.

Lats1 Deletion Causes Increased Germ Cell Apoptosis and Follicular Cysts in Mouse Ovaries.

The Hippo signaling pathway is essential for regulating proliferation and apoptosis in mammalian cells. The LATS1 kinase is a core member of the Hippo signaling pathway that phosphorylates and inactivates the transcriptional co-activators YAP1 and WWTR1. Deletion of Lats1 results in low neonate survival and ovarian stromal tumors in surviving adults, but the effects of Lats1 on early follicular development are not understood. Here, the expression of Hippo pathway components including Wwtr1, Stk4, Stk3, Lats2, and Yap1 transcripts were decreased by 50% in mouse ovaries between 2 and 8 days of age while expression was maintained from 8 days to 21 days and after priming with eCG. LATS1, LATS2, and MOB1B were localized to both germ and somatic cells of primordial to antral follicles. Interestingly, YAP1 was predominantly cytoplasmic, whereas WWTR1 was nuclear in oocytes and somatic cells. Deletion of Lats1 caused an increase in germ cell apoptosis from 1.7% in control ovaries to 3.6% in Lats1 mutant ovaries and a 58% and 32% decrease in primordial and activated follicle numbers in cultured mutant ovaries. Surprisingly, there was an increase in Bmp15 but not Gdf9, Figla, Nobox transcripts or the somatic-specific transcripts Amh and Wnt4 in cultured Lats1 mutant ovaries. Last, Lats1 mutant ovaries developed ovarian cysts at a higher frequency (43%) than heterozygous (24%) and control ovaries (8%). Results showed that the Hippo pathway is active in ovarian follicles and that LATS1 is required to maintain the pool of germ cells and primordial follicles.

mRNA expression pattern of gonadotropin receptors in bovine follicular cysts.

Follicular growth and steroidogenesis are dependent on gonadotropin binding to their receptors in granulosa and theca cells of ovarian follicles. The aim of the present study was to evaluate the expression patterns of follicle-stimulating hormone receptor (FSHR) and luteinizing hormone receptor (LHCGR) in ovarian follicular structures from cows with cystic ovarian disease (COD) as compared with those of regularly cycling cows. Relative real-time RT-PCR analysis showed that the expression of FSHR mRNA in granulosa cells was highest in small antral follicles, then decreased significantly as follicles increased in size, and was lowest in cysts. FSHR mRNA was not detected in the theca cells of any follicular category, including cysts. LHCGR mRNA expression in granulosa cells was significantly higher in large antral follicles than in cysts, and not detected in granulosa cells of small and medium antral follicles. In theca cells, the expression level of LHCGR mRNA in medium antral follicles was higher than in small and large antral follicles, whereas that in follicular cysts it was similar to those in small and medium antral follicles, but higher than that in large antral follicles. Our findings provide evidence that there is an altered gonadotropin receptor expression in bovine cystic follicles, and suggest that in conditions characterized by altered ovulation, such as COD, changes in the signaling system of gonadotropins may play a fundamental role in their pathogenesis.

[Ki-67 and matrix metalloproteinase-9 expression in the follicular cyst, keratocystic odontogenic tumor, and ameloblastoma].

An immunohistochemical study using antibodies against Ki-67 protein was conducted, which characterized the proliferative activity of cells and matrix metalloproteinase-9 (MMP-9) in follicular cyst, variants of keratocystic odontogenic tumor (with a preponderance of hyperkeratosis and parakeratosis), and ameloblastoma. The marked proliferative activity of a parabasal cell layer (28.0+/-8.4%) was found in the keratocystic odontogenic tumor with a preponderance of parakeratosis; the proliferative activity of the peripheral layer of ameloblastoma cells was equal to 7.0+/-5.6%. The maximal matrix metalloproteinase-9 expression in the keratocystic odontogenic tumor with a predominance of hyperkeratosis was 1.1 +/-0.9 conventional units (CU) and that in the ameloblastoma was 1.9+/-1.3 CU versus 0.4+/-0.5 CU in the follicular cyst, keratocystic odontogenic tumor with a preponderance of hyperkeratosis. The values of Ki-67 and MMP-9 expression allow one to distinguish benign odontogenic cysts and tumors (follicular cyst and keratocystic odontogenic tumor with a predominance of hyperkeratosis) and odontogenic tumors characterized by an aggressive clinical course (keratocystic odontogenic tumor with a preponderance of parakeratosis and ameloblastoma).

Acid-base parameters and steroid concentrations in pre-ovulatory follicles and plasma of lactating dairy cows with spontaneous and synchronized oestrus or follicular cyst.

The study aimed to compare the acid-base balance and steroid concentrations between follicular fluids (FF) of pre-ovulatory follicles derived from a spontaneous oestrus (SO), synchronized or induced oestrus (IO) and follicular cysts (CYS) and between FF and blood in dairy cows. Forty-two dairy cows were included in this study. The animals were allocated to three groups: SO (n = 23); IO (n = 11) using GnRH at day 0 and day 9 and PGF2 α at day 7; and animals with CYS (n = 10). The follicular fluids (FF) were aspirated from the cyst/pre-ovulatory follicles (∅ ≥ 15 mm) after SO and after second GnRH dose in IO by transvaginal ultrasound-guided ovum pick-up technique. Blood samples (BL) were collected in heparinized vacutainer tubes. The oxygen tension (pO2) in FF of IO was higher (p < 0.05) than in SO and CYS groups. There were negative correlations (p < 0.001, r = -0.89) between FF and blood pO2. The carbon dioxide tension (pCO2) and lactate level in FF of CYS group were higher (p < 0.05) than in SO and IO groups. There were negative correlations (p < 0.01, r = -0.73) between blood and FF pO2. Oestradiol-17ß concentration in pre-ovulatory follicles and plasma of the SO group was higher (p < 0.001) than in IO and CYS groups. Progesterone concentration in pre-ovulatory follicles and plasma of the SO and IO groups was lower (p < 0.01) than in CYS group. Plasma androstenedione concentration in SO and IO groups was higher (p < 0.05) than in CYS group. In conclusion, acid-base parameters, E2 and P4 levels in the follicular fluid of both IO and CYS groups were deviated greatly from the physiological level (disturbances of intrafollicular/intracystic environment), which may affect the quality of both the oocyte and the granulosa cells.

Comparison of cadherin and integrin localization in bovine cystic and healthy follicles.

As stage progresses in the cystic follicle, granulosa cells are lost. We hypothesized that the granulosa and theca interna layers are detached in association with weakened expression of cell adhesion molecules such as cadherin (cell-cell adhesion) and integrin (cell-extracellular matrix adhesion) in cystic follicles. To elucidate this hypothesis, we immunolocalized these molecules in the granulosa and theca interna and compared them between cystic and small healthy follicles. Sections were immunostained with cadherin and integrin ß1 antibodies and their localizations were compared. Cadherin-positive reaction was seen in the cytoplasma of all granulosa cells. No increase in the frequency of cadherin-positive area in the granulosa layers and the intensity of cadherin immunoreaction in the theca interna was detected in cystic follicles compared with healthy ones. A dense immunoreaction product of integrin ß1 was detected in the theca interna in both cystic and healthy follicles. Intensity of integrin ß1 -immuno reaction in the granulosa layers and integrin ß1 -positive area in the theca interna was significantly lower in the cystic follicle than in the healthy follicles. These results suggest that granulosa and theca interna cells are detached while maintaining the cell-cell adhesion, resulting in the consequent loss of these layers from the cystic follicle.

Inhibition of BMP signaling in P-Cadherin positive hair progenitor cells leads to trichofolliculoma-like hair follicle neoplasias.

BACKGROUND: Skin stem cells contribute to all three major lineages of epidermal appendages, i.e., the epidermis, the hair follicle, and the sebaceous gland. In hair follicles, highly proliferative committed progenitor cells, called matrix cells, are located at the base of the follicle in the hair bulb. The differentiation of these early progenitor cells leads to specification of a central hair shaft surrounded by an inner root sheath (IRS) and a companion layer. Multiple signaling molecules, including bone morphogenetic proteins (BMPs), have been implicated in this process. METHODS: To further probe the contribution of BMP signaling to hair follicle development and maintenance we employed a transgenic mouse that expresses the BMP inhibitor, Noggin, to disrupt BMP signaling specifically in subset of hair follicle progenitors under the control of neuron specific enolase (Nse) promoter. We then studied the skin tumor phenotypes of the transgenic mice through histology, immunohistochemistry and Western Blotting to delineate the underlying mechanisms. Double transgenic mice expressing BMP as well as noggin under control of the Nse promoter were used to rescue the skin tumor phenotypes. RESULTS: We found that the transgene is expressed specifically in a subpopulation of P-cadherin positive progenitor cells in Nse-Noggin mice. Blocking BMP signaling in this cell population led to benign hair follicle-derived neoplasias resembling human trichofolliculomas, associated with down-regulation of E-cadherin expression and dynamic regulation of CD44. CONCLUSIONS: These observations further define a critical role for BMP signaling in maintaining the homeostasis of hair follicles, and suggest that dysregulation of BMP signaling in hair follicle progenitors may contribute to human trichofolliculoma.

Biochemical and hormonal composition of follicular cysts in water buffalo (Bubalus bubalis).

The objective of this study was to examine the follicular fluid biochemical and hormonal changes associated with ovarian follicular cysts in buffalo. Follicular fluid was aspirated from eight cysts and eight preovulatory follicles, and subjected to biochemical and hormonal analyses. Cysts were characterized by a greater (P<0.01) concentration of nitric oxide and lesser concentrations of ascorbic acid and glucose than that of preovulatory follicles (P<0.01 and P<0.05, respectively). Furthermore, follicular cysts had greater concentrations of progesterone (P<0.001), triiodothyronine (T(3)) and cortisol (P<0.05) and lesser concentrations of insulin (P<0.001) than preovulatory follicles. The results indicate follicular cysts in buffalo have an altered biochemical and hormonal composition. The alterations include increases in nitric oxide, progesterone, cortisol and T(3) concentrations with a concurrent reduction in ascorbic acid, insulin and glucose concentrations. The study suggests that greater progesterone concentrations possibly inhibit the onset of LH surge resulting in formation of follicular cysts in buffalo. In addition, it implies the plausible role of intra-ovarian regulators such as nitric oxide, ascorbic acid and insulin in development of the condition.

New insights into the pathogenesis of cystic follicles in cattle: microarray analysis of gene expression in granulosa cells.

Ovarian follicular growth and development are regulated by extraovarian and intraovarian factors, which influence granulosa cell proliferation and differentiation. However, the molecular mechanisms that drive follicular growth are not completely understood. Ovarian follicular cysts are one of the most common causes of reproductive failure in dairy cattle. Nevertheless, the primary cause of cyst formation has not been clearly established. A gene expression comparison may aid in elucidating the causes of ovarian cyst disease. Our objective was to identify differentially expressed genes in ovarian granulosa cells between normal dominant and cystic follicles of cattle. Granulosa cells and follicular fluid were isolated from dominant and cystic follicles collected via either ultrasound-guided aspiration from dairy cows (n = 24) or slaughterhouse ovaries from beef cows (n = 23). Hormonal analysis for progesterone, estradiol, and androstenedione in follicular fluid was performed by RIA. Total RNA was extracted and hybridized to 6 Affymetrix GeneChip Bovine Genome Arrays (Affymetrix, Santa Clara, CA). Abundance of mRNA for differentially expressed selected genes was determined through quantitative real-time reverse-transcription PCR. Follicular cysts showed greater (P < 0.05) progesterone, lesser (P < 0.05) estradiol, and no differences (P > 0.10) in androstenedione concentrations compared with noncystic follicles. A total of 163 gene sequences were differentially expressed (P < 0.01), with 19 upregulated and 144 downregulated. From selected target genes, quantitative real-time reverse-transcription PCR confirmed angiogenin, PGE(2) receptor 4, and G-protein coupled receptor 34 genes as upregulated in cystic follicles, and Indian hedgehog protein precursor and secreted frizzled-related protein 4 genes as downregulated in cystic follicles. Further research is required to elucidate the role of these factors in follicular development and cyst formation.

Bilateral follicular cysts in a water buffalo.

The present short communication puts on record a case of bilateral, multiple follicular cysts in a water buffalo along with a detailed description of its ovarian biometry and follicular fluid composition. The ovarian weight and biometrical parameters were much higher than in normal cycling buffaloes. A total of three follicular cysts were observed, two on the right ovary and one on the left ovary, measuring 4.9, 3.0 and 2.6 cm yielding 21, 9 and 5 ml of follicular fluid, respectively. The cystic fluid was deep yellow in colour with a viscous consistency. The follicular fluid concentrations of glucose, total protein, cholesterol, acid phosphatase, calcium, phosphorus and progesterone in all the cysts were within the range reported previously in normal buffalo follicular fluid; however, the alkaline phosphatase concentration in cyst 1 and total bilirubin concentration in cysts 1 and 2 were higher than the values in normal follicular fluid. In contrast, the levels of urea nitrogen in cysts 1 and 3, and oestradiol in cyst 3 were lower than the normal values. All the three follicles had an oestradiol to progesterone ratio less than 1. The results of our study suggest that follicular cysts in buffalo are oestrogenically inactive and have an altered concentration of certain biochemical and hormonal constituents.

The cytobiological differences between two odontogenic cyst-lining keratinocytes.

PURPOSE: Odontogenic cysts are classified into a developmental group, including follicular cysts (FC) and keratocysts, and an inflammatory group including radicular cysts (RC). In clinical cases, we frequently encounter RC and FC. The purpose of this study was to investigate the cytobiological differences between two odontogenic cyst-lining keratinocytes using a cytobiological approach from the aspect of metabolic function and the degree of maturation of the epithelium. MATERIALS AND METHODS: Samples of odontogenic cyst-lining keratinocytes and oral keratinocytes collected at surgery, and of cultured oral keratinocytes, were analyzed (1) by immunohistochemical staining of granulocyte macrophage colony stimulating factor (GM-CSF), human beta defensin-2 (HBD-2) and chemokine receptor 6 (CCR6) expressing cell (Langerhans cell, helper T cell and suppressor T cell) antibodies, (2) by reverse transcription-polymerase chain reaction (RT-PCR) to determine the expression of GM-CSF and HBD-2 mRNA and (3) by gas chromatography to evaluate the composition of fatty acids (16:0, 18:2, 20:4) in the cell membranes of the keratinocytes. RESULTS: 1. Immunohistochemical staining indicated that HBD-2 and GM-CSF expression were higher in RC than in FC. 2. The same results were obtained from the RT-PCR analysis. 3. The % composition of palmitic acid (16:0) was significantly higher in the RC-lining keratinocytes (38.62±5.86%) and in the FC-lining keratinocytes (30.37±1.38%) than in the normal gingiva (23.00±1.40%). The % composition of essential fatty acids (18:2+20:4) was significantly higher in the FC-lining keratinocytes (26.20±3.55%) than in the RC-lining keratinocytes (20.50±8.17%). CONCLUSION: The present study demonstrated definite cytobiological evidence of the differences between RC and FC.

Isosexual pseudoprecocious puberty in a 2(1/2)-year-old girl presenting with intense skin pigmentation.

BACKGROUND: Ovarian follicular cyst producing estradiol is a rare cause of isosexual pseudoprecocious puberty. Intense pigmentation of breast papillae, areolae, and labia minora is also rarely reported in the literature. CASE: We describe a 2(1/2) year old girl presenting with signs of precocious puberty and advanced bone age due to a large follicular cyst. Estradiol and Dehydro-epiandrosterone sulfate (DHEAS) levels were remarkably elevated. Hyperpigmentation was also noted. Salpingoophorectomy resulted in regression of precocity and depigmentation, but DHEAS serum levels remained elevated. SUMMARY AND CONCLUSION: High levels of circulating estradiol due to an ovarian follicle can induce precocious puberty and pigmentation of the skin which regresses after surgical removal of the cyst. Elevated DHEAS levels may be the initiating event causing the formation of the large follicular cyst.

Reactive oxygen species (ROS): involvement in bovine follicular cysts etiopathogenesis.

Ovulation is compared to an acute inflammatory process during which vasoactive agents, prostanoids, leukotrienes and Reactive Oxygen Species (ROS) develop. The aim of this study was to evaluate the levels of ROS in cystic and follicular fluid, in order to establish their involvement in the etiopathogenesis of Cystic Ovarian Follicle (COF) in dairy cows. The study was conducted in 30 healthy cows (group C) and 30 cows affected by COF (group COF). The fluid of follicular cysts and of preovulatory follicles was drawn by means of ultrasound guided aspiration from the cows of both groups. The fluid obtained was analyzed by a photometric analytical system to detect ROS level. ROS concentration was statistically lower in the cystic fluid than in the follicular one (62.4 +/- 13.36 U.Carr vs. 84.89 +/- 26.99 U.Carr) (p<0.05), thus suggesting that an alteration of the cascade responsible for ROS production may be implicated in the complex etipathogenesis of COF.